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Related Experiment Videos

A permeabilized cell system that assembles filamentous bacteriophage

J N Feng1, M Russel, P Model

  • 1Laboratory of Genetics, The Rockefeller University, New York, NY 10021, USA.

Proceedings of the National Academy of Sciences of the United States of America
|April 15, 1997
PubMed
Summary

A new in vitro system allows filamentous phage assembly using exogenous thioredoxin. This system requires functional thioredoxin, DNA packaging signals, and protein pIV, demonstrating ATP hydrolysis and proton motive force are crucial for assembly.

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Area of Science:

  • Molecular Biology
  • Virology
  • Biochemistry

Background:

  • Filamentous phage assembly is a complex process occurring in vivo.
  • Understanding the in vitro requirements for phage assembly can elucidate the underlying molecular mechanisms.

Purpose of the Study:

  • To develop a permeabilized cell system for in vitro filamentous phage assembly.
  • To investigate the energy requirements for filamentous phage assembly.

Main Methods:

  • Developed a permeabilized cell system for in vitro phage assembly.
  • Added exogenous thioredoxin to the system.
  • Assessed the impact of ATP hydrolysis and proton motive force on assembly.

Main Results:

  • The in vitro system requires functional thioredoxin, intact packaging signals on DNA, and the assembly protein pIV.

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  • The system is insensitive to inhibitors of protein or DNA synthesis.
  • Filamentous phage assembly in this system requires ATP hydrolysis and is influenced by proton motive force.

    • Conclusions:

    • A functional in vitro system for filamentous phage assembly has been established.
    • The in vitro system recapitulates the in vivo component requirements.
    • Filamentous phage assembly is an energy-dependent process involving ATP hydrolysis and proton motive force.